Ricin toxin ricinus communis agglutinin 2

Ricin toxin (Ricinus communis agglutinin II) was purchased from Vector Laboratories (Burlingame, CA). Ricin was dialyzed against phosphate-buffered saline (PBS) at 4°C in 10,000-molecular-weight (MW)-cutoff Slide-A-Lyzer dialysis cassettes (Pierce, Rockford, IL), prior to use in cytotoxicity studies. Fetal calf serum was purchased from Gibco-Invitrogen (Carlsbad, CA). Cells were maintained in a humidified incubator at 37°C with 5% CO₂. Recombinant human tumor necrosis factor alpha (TNF-α), recombinant human soluble TRAIL (sTRAIL)/Apo2L/CD253, anti-human sTRAIL-(s)-Apo2L, and rabbit polyclonal anti-TRAIL antibody were purchased from Peprotech (Rocky Hill, NJ). Human TNF-α neutralizing rabbit Ab was purchased from Cell Signaling Technology (Danvers, MA). Z-LEHD-FMK, Z-VAD-FMK, Z-DEVD-FMK, and Z-IETD-FMK were purchased from ApexbBio (Taiwan). Necrostatin-1 (Nec-1), GSK′872, and necrosulfonamide (NSA) were purchased from EMD Millipore (Burlington, MA). Unless noted otherwise, all other chemicals were obtained from MilliporeSigma (St. Louis, MO). Murine MAbs against ricin toxin’s A subunit (PB10, SyH7, GD12, and IB2) and B subunit (24B11, SylH3, MH3, 8A1, 8B3, LF1, and LC5) were purified by protein A chromatography at the Dana Farber Cancer Institute (DFCI) Monoclonal Antibody Core facility (Boston, MA), as described previously (36 (link), 55 (link)).

Ricin toxin (Ricinus communis agglutinin II) and ricin-FITC were purchased from Vector Laboratories (Burlingame, CA). Tween-20, Triton X-100, and Hoechst 33342 were purchased from Sigma-Aldrich (St. Louis, MO). Commercial secondary and primary Abs are described in Supplementary Table 2. Glass coverslips (22 × 22 mm square, 1.5 mm) were purchased from Corning-Fischer Scientific (Suwanee, GA). Tissue culture-treated dishes (35 mm x 15 mm) were purchased from CELLTREAT Scientific Products (Shirley, MA). Cytofix/Cytoperm™ Fixation/Permeabilization Solution was purchased from BD Biosciences (San Jose, California). Ammonium chloride was purchased from Sigma-Aldrich. HeLa and Vero cells were purchased from the American Type Culture Collection (Manassas, VA). Unless otherwise noted, all cell lines were maintained in a humidified incubator at 37 °C with a 5% CO₂ atmosphere.